ADAMs (a disintegrin and metalloprotease) or MDCs (metalloprotease disintegrin cysteine-rich proteins) form a family of type I transmembrane proteins. Owing to their multidomain structure consisting of pro-, metalloprotease, disintegrin-like, cystein-rich, EGF-like, transmembrane and cytoplasmic domains, ADAMs are capable of four physiological functions: cell adhesion, cell fusion, cell signalling and proteolysis.
ADAMs are implicated in physiological processes such as fertilization, myogenesis and neurogenesis, and are also involved in a number of pathological processes by releasing cytokines and their receptors under inflammatory conditions (see for example Moss and Bartsch, 2004). To date, 40 members are known in different species (see table of ADAMs at http colon-slash-slash www.people.virginia.edu/;jw7g/). Approximately half of these ADAMs contain the catalytic consensus sequence HEXXHHXXGXXHD (SEQ ID NO:1) in their metalloprotease domains and are therefore predicted to be catalytically active. Proteolysis of membrane-anchored precursor proteins is a key event in signalling cascades (Blobel, 2005) and this process has been termed ectodomain shedding (Peschon et al., 1998). A number of ADAM substrates have been defined either by their physiological role or by their cellular localisation (Seals and Courtneidge, 2003; Moss and Bartsch, 2004; Blobel, 2005). So far, no consensus sequence on the substrate side has been reported, making prediction of potential cleavage sites difficult. Rather, ADAMs recognise a structural footprint on the extracellular part of the membrane protein, and it is assumed that cleavage occurs in the juxtamembraneous regions of these membrane proteins.
One particular area of research interest is the ADAM family member ADAM8, originally identified as MS2 or CD156 on mouse macrophages (Yoshida et al., 1990). ADAM8 is expressed in several tissues, such as thymus, cartilage, bone, brain and spinal cord, during embryonic development. As embryonic development of ADAM8-deficient mice appears normal, ADAM8 does not seem to be essential for developmental processes (Kelly et al., 2005). Rather, it is more likely that ADAM8 has a specific function in cytokine response which is reflected by its distinct expression in immune cells like B-cells, leukocytes, neutrophils, macrophages, and dendritic cells. In response to inflammatory stimuli such as lipopolysaccharide (LPS) and tumour necrosis factor a (TNF-α), ADAM8 expression is upregulated in most immune cells. In addition, ADAM8 is upregulated in the central nervous system (CNS) under inflammatory conditions in activated glia cells—astrocytes and microglia—indicating its involvement in neuron-glia signalling, in particular in neuroinflammatory disorders (Schlomann et al., 2000). Similarly to glial cells, ADAM8 is expressed in neurons at low levels, and expression is induced by TNF-α, suggesting that under inflammatory conditions in the CNS, neuronal ADAM8 activity is significantly increased (Schlomann et al., 2000).
In the lung, ADAM8 is upregulated under experimental induction of allergic asthma by inflammatory cytokines (King et al., 2004), suggesting that ADAM8 plays a role in the pathogenesis of allergic asthma. The induction of experimental asthma in ADAM8 deficient mice almost suppresses the asthmatic phenotype, arguing for an essential role of ADAM8 in the pathogenesis of allergic asthma.
All these findings underline the importance of ADAM8 in inflammatory processes and the necessity to inhibit ADAM8 activity under those pathological conditions where ADAM8 activity has detrimental effects.
Recent attention has focussed on ADAM8 as a molecule upregulated in various tumours. ADAM8 is considered a prognostic marker for lung adenocarcinomas (Ishikawa et al., 2004) and renal cell carcinomas (Roemer et al., 2004). In addition, in brain tumours such as glioblastoma, oligoastrocytoma, and ependymoma, ADAM8 expression was increased compared to normal brain controls, and ADAM8 expression in glioblastoma correlates with malignancy and invasive activity (Wildeboer et al., 2006). In B-cells, ADAM8 cleaves the low-affinity IgE receptor (CD23, Fc′RII) suggesting a role in immune modulation (Fourie et al., 2003). Thus ADAM8 is implicated in allergy and inflammation by ectodomain shedding. A number of substrates of ADAM8 and their corresponding cleavage sites are known (Amour et al., 2000; Schlomann et al., 2002; Fourie et al., 2003; Naus et al., 2004, Naus et al., 2006).
Accordingly, inhibition of ADAM proteases is an attractive therapeutic target for many diseases. For example, WO 01/09189 discloses methods for diagnosing or treating neoplastic cell growth (e.g. certain cancers) involving agents which target ADAM8. WO 2005/090991 discloses methods for diagnosing or treating non-small cell lung cancer using agents such an ADAM8 siRNA that reduces expression of the ADAM8 gene. WO 2004/024089 discloses treating diseases associated with pathological neovascularisation (such as cancer, diabetic retinopathy and psoriasis) using agents which inhibit ADAM9 or ADAM15. WO 01/94377 discloses that ADAM 8 is also a therapeutic target for the treatment of allergy and asthma, whereas ADAM15 is a target for the treatment of osteoarthritis and atherosclerosis. The TNF-α-converting enzyme TACE (ADAM17) is a target for the treatment of inflammatory diseases including rheumatoid arthritis and ulcerative colitis (WO 97/35538 and Sekut et al. 1998, Expert Opin. Invest. Drugs. 7:1825-1839).
However, a number of the methods which have been identified to date for inhibiting metalloproteases such as ADAMs suffer from serious drawbacks. For instance, hydroxamate compounds such as marimastat and batimastat (BB-94) are typically broad-spectrum inhibitors and suffer from a lack of specificity for particular ADAMs. Consequently these compounds have shown deleterious side effects in clinical trials, which seriously restricts their usefulness as therapeutic agents. Therefore there is still a need for improved and more specific methods and reagents for inhibiting ADAM proteases, in particular for treating diseases such as cancer, inflammatory conditions and allergy.